RNA forms complex tertiary structures that impart diverse functions [1,2]. For example, RNA catalyzes reactions [3], regulates gene expression [4, 5], encodes protein, and plays other essential roles in biology. Therefore, RNA is an interesting and important target for developing drugs or probes of function [6,7]. It is a vastly underutilized target, however, mainly because of the limited information available on RNA ligand interactions that could facilitate rational design.
One advantage of using RNA as a drug target is that secondary structure information, which includes the motifs that comprise an RNA, can be easily obtained from sequence by free energy minimization [8,9] or phylogenic comparison [10]. RNA tertiary structures are composites of the secondary structural motifs and the long-range contacts that form between them. Furthermore, RNA motifs can have similar properties both as isolated systems and as parts of larger RNAs. For example, aminoglycoside antibiotics affect the structure of the bacterial rRNA A-site similarly when they bind the entire ribosome or an oligonucleotide mimic of the bacterial rRNA A-site [11-16]. Studies on the binding of aminoglycosides and streptamine dimers to RNA hairpins [17-20] have facilitated the development of compounds to combat multidrug resistance by causing plasmid incompatibility [19, 20]. These results show that the identification of RNA motifs that bind small molecules can be useful for targeting the larger RNAs that contain them.
However, since RNA can adopt diverse structures, internal and hairpin loops for example, an understanding how to target RNA with small molecules and other ligands has not yet been achieved. Current methods to study and identify RNA ligand interactions include systematic evolution of ligands by exponential enrichment (“SELEX”) [21, 22], structure-activity relation-ships (“SAR”) by mass spectrometry (“MS”) [23-26] and NMR [27], and chemical microarrays [28-30]. These methods probe RNA space (SELEX) or chemical space (SAR by MS and NMR and chemical microarrays) separately. However, these methods do not permit a systematic study of RNA-ligand interactions.
For all of the above reasons, a need exists for methods that can be used to systematically identify RNA-ligand interactions, and the present invention is directed, in part, to addressing this need.